The objective of this study is to elucidate the intracellular interactions of the HIV regulatory protein Nef with host cell activities. Early in the HIV-1 infectivity process this retrovirus expresses regulatory proteins, with the Nef transcript representing nearly 80% of total viral mRNA. In vivo infectivity by the closely related simian form of HIV is achieved in the absence of Nef expression, but there is an absolute requirement for Nef in the development of the immunodeficiency. This essential activity of the Nef protein in the development of AIDS has not been defined. We had previously found that Nef expression independently cause enhancement of T cell activation and modulation the CD4 molecule, the HIV receptor. It was also shown that Nef expression specifically inactivated inositol triphosphate receptor function. Preliminary work suggests that Nef expression can also alter monocyte function; induction of IL-6, but not IL-10, is enhanced in a monocytic cell line. To determine the nature of Nef molecular interactions that lead to these signal pathway alterations, we have constructed a Nef fusion protein that can be expressed in cultured cell lines and purified from cell lysates on an affinity column. This system has demonstrated that in the presence of detergent the Nef protein is associated with a serine/threonine kinase. This kinase is expressed in T cell and has the capacity to autophosphroylate.